| Chinese dwarf cherry(Cerasus humilis)is a unique shrubby fruit tree resource in China,which is characterized by high nutrition and high resistance,as well as enormous potential and economic values.Significant variation in acidity among different germplasms and lack of low-acid varieties make the breeding of varieties with low acidity an important goal.In this study,the fruits of high-acid Chinese dwarf cherry variety ’Nongda4’,low-acid genotype’DS-1’ and their hybrid progenies(F1 population)were selected as test materials.Firstly,determination of the organic acid contents and transcriptome sequencing were conducted in fruits of ’Nongda4’ and ’DS-1’ at different developmental stages;Secondly,determination of organic acid contents and whole genome re-sequencing of the F1 population were also carried out.Based on transcriptome analysis and QTL mapping techniques,key genes related to organic acid metabolism in Chinese dwarf cherry fruits were screened,and functional verification was then conducted.The results of this study will provide theoretical basis for revealing the genetic mechanism and regulation of acidity of Chinese dwarf cherry fruit.The main research results are as follows:1.The genetic variation of fruit traits of ’Nongda4×DS-1’ hybrid population for two consecutive years was analyzed.The fruit titratable acid content,soluble solids,single fruit weight,nuclear weight,transverse diameter and longitudinal diameter of the hybrid population followed continuous normal distribution patterns,indicating these fruit traits were typical quantitative traits that were controlled by multiple genes;The variation coefficient and genetic transmission of fruit titratable acid content of hybrid progenies were relatively high,and the titratable acid content was significantly correlated with soluble solids content,but having no correlation with other traits,therefore,the genetic mechanism of organic acid accumulation was relatively independent.2.The organic acid content and transcriptome of fruit samples at different developmental stages of the high acid variety ’Nongda4’ and low acid genotype ’DS-1’ of Chinese dwarf cherry were determined.The titratable acid content of both varieties exhibited an increasing-then-decreasing trend during fruit development,and the organic acid content of’Nongda4’ always significantly higher than that of ’DS-1’.According to the deferentially expressed genes(DEGs)related to organic acid metabolism in the transcriptome and the WGCNA method,MSTRG.4608(t DT),MSTRG.15150 and MSTRG.1941(MDH),MSTRG.276 and MSTRG.5073(VHA),MSTRG.13863(ME),MSTRG.20567(PEPCK)correlating to the acidity of Chinese dwarf cherry fruit were selected and their complex regulatory networks were constructed;the q RT-PCR analysis of 36 Chinese dwarf cherry germplasm further verified that these 7 key genes were significantly correlated with acidity accumulation of Chinese dwarf cherry fruit.3.Whole genome re-sequencing of 208 progenies of the F1 population and their parents were conducted.A large number of SNP markers were screened and a high density genetic map of Chinese dwarf cherry were constructed accordingly.The genetic map contained 8linked groups and 5371 SNP markers.The total genetic distance was 1321.5 c M,with an average distance of 0.26 c M between markers.Based on the phenotypic data of titrable acid content in fruits of ’Nongda4’ × ’DS-1’ F1 population,QTL mapping was performed by compound interval mapping.A total of 13 QTLs loci related to the titratable acid content of the fruits were identified,mainly located on LG1,LG2,LG3,LG6 and LG7,with a phenotypic variation contribution rate from 7.1 to 13.1.One structural gene(VHA-e1,LG3)and two transcription factors(MYB44 and b HLH148,LG1)significantly related to fruit acidity were screened from the,providing a direction for further research on the regulatory mechanism of fruit acidity in Chinese dwarf cherry.4.The MDH and VHA-G of the key genes of organic acid metabolism in Chinese dwarf cherry fruit were homologously cloned,and their roles in organic acid metabolism were analyzed by transient overexpression and silencing methods.The results showed that after overexpression of the MDH and VHA-G genes,the titratable acid and malic acid contents were significantly increased compared to the control group(P<0.01).After silencing the MDH and VHA-G genes,the titratable acid and malic acid contents were significantly decreased compared to the control group(P<0.01).These results suggest that MDH and VHA-G play a regulatory role in organic acid accumulation in Chinese dwarf cherry fruit,providing a theoretical basis for further studies on the mechanism of these genes in the development of Chinese dwarf cherry fruit. |
PDF Full Text Request